Articular cartilage, an avascular tissue found at the ends of articulating bones, has limited natural capacity to heal. During normal cartilage ontogeny, mesenchymal stem cells condense to form areas of high density and proceed through a series of developmental stages that ends in the mature chondrocyte. The final hyaline cartilage tissue contains only chondrocytes that are surrounded by a matrix composed of type II collagen, sulfated proteoglycans, and additional proteins. The matrix is heterogenous in structure and consists of three morphologically distinct zones: superficial, intermediate, and deep. Zones differ among collagen and proteoglycan distribution, calcification, orientation of collagen fibrils, and the positioning and alignment of chondrocytes (Archer et al., 1996, J. Anat. 189(1):23-35; Morrison et al., 1996, J. Anat. 189(1): 9-22; and Mow et al., 1992, Biomaterials 13(2): 67-97). These properties provide the unique mechanical and physical parameters to hyaline cartilage tissue.
The meniscus, a C-shaped cartilaginous tissue, performs several functions in the knee including load transmission from the femur to the tibia, stabilization in the anterior-posterior position during flexion, and joint lubrication. Damage to the meniscus results in reduced knee stability and knee locking. Over 20 years ago, meniscectomies were performed which permitted immediate pain relief, but were subsequently found to induce the early onset of osteoarthritis (Fairbank, J. Bone Joint Surg. 30B: 664-670; Allen et al., 1984, J. Bone Joint Surg. 66B:666-671; and Roos et al., 1998, Arth. Rheum. 41:687-693). More recently, partial meniscectomies and repair of meniscal tears have been performed (FIGS. 9A-D; Jackson, D., ed., 1995, Reconstructive Knee Surgery Master Techniques in Orthopedic Surgery, ed. R. Thompson, Raven Press: New York). However, partial resection results in the loss of functional meniscus tissue and the early onset of osteoarthritis (Lynch et al., 1983, Clin. Orthop. 172:148-153; Cox et al., 1975, Clin. Orthop. 109:178-183; King, 1995, J. Bone Joint Surg. 77B:836-837). Additionally, repair of meniscal tears is limited to tears in the vascular ⅓ of the meniscus; tears in the semivascular to avascular ⅔ are not repairable (FIGS. 9A-D; Jackson, ibid.). Of the approximately, 560,000 meniscal injuries that occur annually in the United States, an estimated 80% of tears are located in the avascular, irreparable zone. Clearly, a method that both repairs “non-repairable” tears or that can induce regeneration of resected menisci would be valuable for painless musculoskeletal movement and prevention of the early onset of osteoarthritis in a large segment of the population.
The proximal, concave surface of the meniscus contacts the femoral condyle and the distal, flat surface contacts the tibial plateaus. The outer one-third of the meniscus is highly vascularized and contains dense, enervated, connective tissue. In contrast, the remaining meniscus is semivascular or avascular, aneural tissue consisting of fibrochondrocytes surrounded by abundant extracellular matrix (McDevitt et al., Clin. Orthop. Rel. Res. 252:8-17). Fibrochondrocytes are distinctive in both appearance and function compared to undifferentiated fibroblasts. Fibroblasts are elongated cells containing many cellular processes and produce predominantly type I collagen. The matrix produced by fibroblasts does not produce a sufficient mechanical load. In contrast, fibrochondrocytes produce type I and type II collagen and proteoglycans. These matrix components support compressive forces that are commonly exerted on the meniscus during musculoskeletal movement.
In the 1960's, demineralized bone matrix was observed to induce the formation of new cartilage and bone when implanted in ectopic sites (Urist, 1965, Science 150:893-899). The components responsible for the osteoinductive activities were termed Bone Morphogenetic Proteins (BMP). At least seven individual BMP proteins were subsequently identified from bone (BMP 1-7) and amino acid analysis revealed that six of the seven BMPs were related to each other and to other members of the TGF-β superfamily. During endochondral bone formation, TGF-β family members direct a cascade of events that includes chemotaxis, differentiation of pluripotential cells to the cartilage lineage, maturation of chondrocytes to the hypertrophic stage, mineralization of cartilage, replacement of cartilage with bone cells, and the formation of a calcified matrix (Reddi, 1997, Cytokine & Growth Factor Reviews 8:11-20). Although individual, recombinant BMPs can induce these events, the prevalence of multiple TGF-β family members in bone tissue underlies the complexity involved in natural osteogenesis.
Bone Protein (Sulzer Orthopedics Biologics, Wheatridge, Colo.), also referred to herein as BP, is a naturally derived mixture of proteins isolated from demineralized bovine bones that has osteogenic activity in vitro and in vivo. In the rodent ectopic model, BP induces endochondral bone formation or bone formation through a cartilage intermediate (Damien, C. et al., 1990, J. Biomed. Mater. Res. 24:639-654). BP in combination with calcium carbonate promotes bone formation in the body (Poser and Benedict, PCT Publication No. WO95/13767). In vitro, BP has been shown to promote differentiation to cartilage of murine embryonic mesenchymal stem cells (Atkinson et al., 1996, In “Molecular and Developmental Biology of Cartilage”, Bethesda, Md., Annals New York Acad. Sci. 785:206-208; Atkinson et al., 1997, J. Cell. Biochem. 65:325-339) and of adult myoblast and dermal cells (Atkinson et al., 1998, 44th Annual Meeting, Orthopaedic Research Society, abstract). To ensure chondrogenesis in these in vitro systems, however, culture conditions must be tightly controlled throughout the culture period, including by controlling cellular organization within the culture, optimizing media formulations, and adding exogenous factors that must be carefully established to maximize chondrogenesis over mitogenesis. Such optimization of conditions makes the application of the disclosed in vitro methods to an in vivo system unrealistic and unpredictable. In addition, although in vitro cultures of adult myoblast and dermal cells initially resulted in chondrogenesis, the effect was only transient and over time, the cultures reverted to their original phenotype. Although certain embryonic and precursor cell types showed prolonged chrondrogenesis in vitro in these studies, it would be unpredictable or even impossible in the case of embryonic cells that these specific cell types could be recruited to a site in vivo in an adult patient.
Atkinson et al., in PCT Application No. PCT/EP/05100, incorporated herein by reference in its entirety, describe a delivery system for osteoinductive and/or chondroinductive mixture of naturally derived factors for the induction of cartilage repair.
Hunziker (U.S. Pat. Nos. 5,368,858 and 5,206,023) describes a cartilage repair composition consisting of a biodegradable matrix, a proliferation and/or chemotactic agent, and a transforming factor. A two-stage approach is used where each component has a specific function over time. First, a specific concentration of proliferation/chemotactic agent fills the defect with repair cells. Second, a larger transforming factor concentration, preferably provided in conjunction with a delivery system, transforms repair cells to chondrocytes. The second stage delivery of a high concentration of transforming factor in a delivery system (i.e., liposomes) was required to obtain formation of hyaline cartilage tissue at the treatment site.
Chen and Jeffries (U.S. Pat. No. 5,707,962) describe osteogenic compositions consisting of collagen and sorbed factors to enhance osteogenesis.
Valee and King (U.S. Pat. No. 4,952,404) describe healing of injured, avascular meniscus tissue by release of the angiogenic factor, angiogenin, over at least 3 weeks.
Previously, Amoczky et al. described a method using an autogenous fibrin clot to repair an avascular, circular lesion in canine menisci (Amoczky et al., 1988, J. Bone Joint Surg. 70A:1209-1217). This approach enhanced repair of meniscal tissue compared to controls lacking the fibrin clot. However, the repair tissue was not meniscus-like tissue, but rather connective scar tissue.
Hashimoto et al. described a method using fibrin sealant with or without endothelial cell growth factor in avascular, circular meniscal defects in the canine model (Hashimoto et al., 1992, Am. J. Sports Med. 20:537-541). The growth factor added a modest benefit compared to healing with fibrin sealant alone and this additional effect was not observed until three months after treatment, indicating an indirect contribution of the growth factor. In addition, the defect was filled with hyaline cartilage-like cells, which are not typically present in normal meniscus tissue.
Shirakura, et al. describe the use of an autogenous synovium graft sutured into meniscal tears. While the synovium did enhance healing in ⅓ of the animals, the grafts healed with fibrous tissue, not fibrocartilaginous tissue normally observed in meniscus tissue (Shirakura, 1997, Acta. Orthop. Scand. 68:51-54). Furthermore, ⅔ of the grafts did not heal.
The molecular mechanism for cartilage and bone formation has been partially elucidated. Both bone morphogenetic proteins (BMP) and transforming growth factor β (TGFβ) molecules bind to cell surface receptors (i.e., TGFβ/BMP receptors) to initiate a cascade of signals to the nucleus that promotes proliferation, differentiation to cartilage, and/or differentiation to bone (Massague, 1996, Cell 85:947-950). In 1984, Urist described a substantially pure, but not recombinant, BMP combined with a biodegradable poly (lactic acid) polymer delivery system for bone repair (U.S. Pat. No. 4,563,489). This system blends together equal quantities of BMP and poly(lactic acid) (PLA) powder (100 μg of each) and decreases the amount of BMP required to promote bone repair.
Hattersley et al. (WO 96/39170) disclose a two factor composition for inducing cartilaginous tissue formation using a cartilage formation-inducing protein and a cartilage maintenance-inducing protein. Specific recombinant cartilage inducing proteins are specified as BMP-13, MP-52 and BMP-12, and specific cartilage maintenance-inducing proteins are specified as BMP-9. In one embodiment, BMP-9 is encapsulated in a resorbable polymer system and delivered to coincide with the presence of cartilage formation inducing protein(s).
Laurencin et al. (U.S. Pat. No. 5,629,009) disclose a chondrogenesis-inducing device, consisting of apolyanhydride and polyorthoester, that delivers water soluble proteins derived from demineralized bone matrix, TGFβ, epidermal growth factor (EGF), fibroblast growth factor (FGF) or platelet-derived growth factor (PDGF).
Bentz et al. (PCT Publication No. WO 92/09697) have described the use of a bone morphogenetic protein (BMP) with a TGFβ protein for bone repair. The ratio of BMP to TGFβ in the mixture is in the range of 10:1 to 1:10. The addition of TGF-β with either BMP-2 or BMP-3 results in increased osteoinductive activity and an increased ratio of cartilage to bone when compared to either factor alone (Bentz et al., Matrix 11:269-275 (1991); Ogawa et al., J. Biol. Chem., 267(20): 14233-7 (1992); WO92/09697). However, this composition produced substantial bone in the rodent subcutaneous assay.
Bulpitt and Aeschlimann found that TGFβ-2 and BMP-2 led to accelerated bone formation and decreased cartilage formation in the rat ectopic bone formation assay (Bulpitt et al., Tissue Engineering, pp. 162-169 (1999)).
Other studies demonstrate no or little effect of the combination of TGFβ-1 or -2 with BMP. In vitro, the combination of TGFβ-1 and porcine BMP demonstrated no synergistic effects on collagen production (Kim et al., Biochem. Mol. Biol. Int'l, 33(2):253-261 (1994)). Similarly, Ballock et al., demonstrated no synergy between TGFβ-1 and BMP-3 for collagen production in periosteum derived cells in vitro (Ballock et al., J. Ortho. Res., 15:463-7 (1997)).
In the Rosen modified Sampath-Reddi rodent assay (Sampath et al., Proc. Nat'l Acad. Sci. USA, 80(21):6591-5 (1983)), BMP-2 containing implants showed cartilage and bone formation after ten days and mostly bone (no cartilage) after 21 days (U.S. Pat. No. 5,658,882).
Previously, Li and Stone (U.S. Pat. No. 5,681,353) have described a Meniscal Augmentation Device that consists of biocompatible and bioresorbable fibers that acts as a scaffold for the ingrowth of meniscal fibrochondrocytes, supports normal meniscal loads, and has an outer surface that approximates the natural meniscus contour. After partial resection of the meniscus to the vascular zone, this device is implanted into the resulting segmental defect. The results have been described in both canines and humans (Stone et al., 1992, Am. J. Sports Med. 20:104-111; and Stone et al., 1997, J. Bone Joint Surg. 79:1770-1777).
The Meniscus Augmentation Device, the research reports and patent disclosures described above, and current repair surgeries provide encouraging results in the area of cartilage repair, but are not satisfactory to induce repair of “non-repairable” avascular tears in which the repair tissue is meniscus tissue, and are not satisfactory to produce short patient rehabilitation times and regenerated meniscus tissue in the vascular zone. Furthermore, no reports have been described that demonstrate enhanced healing rates of “repairable” meniscal tears in vivo.